Betaine-assisted multiplex recombinase polymerase amplification coupled lateral flow assay for simultaneous detection and typing of variants in human respiratory virus

Journal article

Authors/Editors

PATSAMON RIJIRAVANICH

Strategic Research Themes

Medical Diagnostics (Smart Healthcare)

Publication Details

Author list: Karunaithas, S.; Chaibun, T.; Rijiravanich, P.; Puenpa, J.; Yong Poovorawan, Y.; Lee, L.S.; Promptmas, C.; Athamanolap, P.; Lertanantawong, B.

Publisher: Elsevier

Publication year: 2026

Journal: Sensors and Actuators B: Chemical (0925-4005)

Volume number: 446

Start page: 138742

ISSN: 0925-4005

eISSN: 1873-3077

URL: https://www.scopus.com/inward/record.uri?eid=2-s2.0-105015507213&doi=10.1016%2Fj.snb.2025.138742&partnerID=40&md5=017310bf666c953b212500cc91e8a3bb

Languages: English-Great Britain (EN-GB)

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Abstract

Rapid and accurate diagnosis of infectious diseases is of paramount importance for effective prevention and treatment. The recent COVID-19 pandemic accelerated the development of on-site detection tools for rapid screening and diagnosis. Recombinase polymerase amplification (RPA) is a well-established isothermal amplification method that offers a potential alternative to polymerase chain reaction for point-of-care applications. However, RPA's specificity remains limited due to non-specific amplification. In this study, we developed a betaine-assisted multiplex RPA system coupled with lateral flow assay for the simultaneous detection and typing of human respiratory virus variants, addressing the issue of non-specific amplification. Our results demonstrated that the addition of 8 µL of betaine per reaction effectively eliminated non-specific amplification in the multiplex RPA system. This method was applied to detect SARS-CoV-2 variants, and its analytical and clinical performance was evaluated. The findings revealed that betaine-assisted multiplex RPA specifically detects SARS-CoV-2 variants with a limit of detection of 1 fM, visible to the naked eye. Furthermore, 120 clinical samples, which included negative, alpha variant, and delta variant cases, were tested and showed 100 % concordance with the standard RT-qPCR method. This proof of concept can be adapted for the detection of various pathogens, particularly for screening emerging variants during outbreaks. © 2025 Elsevier B.V.

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